Cell-permeable probe for identification and imaging of sialidases.

Charng-Sheng Tsai, Hsin-Yung Yen, Meng-I Lin, Tsung-I Tsai, Shi-Yun Wang, Wen-I Huang, Tsui-Ling Hsu, Yih-Shyun E Cheng, Jim-Min Fang, Chi-Huey Wong

Index: Proc. Natl. Acad. Sci. U. S. A. 110(7) , 2466-71, (2013)

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Abstract

Alkyne-hinged 3-fluorosialyl fluoride (DFSA) containing an alkyne group was shown to be a mechanism-based target-specific irreversible inhibitor of sialidases. The ester-protected analog DFSA (PDFSA) is a membrane-permeable precursor of DFSA designed to be used in living cells, and it was shown to form covalent adducts with virus, bacteria, and human sialidases. The fluorosialyl-enzyme adduct can be ligated with an azide-annexed biotin via click reaction and detected by the streptavidin-specific reporting signals. Liquid chromatography-mass spectrometry/mass spectrometry analysis on the tryptic peptide fragments indicates that the 3-fluorosialyl moiety modifies tyrosine residues of the sialidases. DFSA was used to demonstrate influenza infection and the diagnosis of the viral susceptibility to the anti-influenza drug oseltamivir acid, whereas PDFSA was used for in situ imaging of the changes of sialidase activity in live cells.